本文選題：藍光 + 抑制　； 參考：《沈陽農(nóng)業(yè)大學》2017年碩士論文

【摘要】：果實的成熟過程是決定其貯藏性的重要因素。研究果實的成熟及調(diào)控機理對于人工調(diào)控果實貯藏性有重要的理論及實踐意義。秋子梨(Pyrus ussuriensis Maxim)是我國北方地區(qū)特有的梨資源,大部分秋子梨包括'南果梨'是呼吸躍變型果實,其成熟過程主要受乙烯調(diào)控。除此之外,光、溫度等外界環(huán)境因素也可以調(diào)控果實的成熟過程。近年來許多研究證明了藍光能夠抑制呼吸躍變型果實成熟過程中的乙烯合成,從而影響果實的貯藏性,但其分子機制尚不清楚。本研究以'南果梨'果實為試材,研究了藍光通過其信號途徑中的轉(zhuǎn)錄因子PuHYH1抑制乙烯合成關(guān)鍵基因PuACS1的表達進而調(diào)控果實成熟過程中的乙烯合成的分子機制。主要結(jié)果如下:1.從梨基因組克隆了光信號轉(zhuǎn)導途徑中的重要轉(zhuǎn)錄因子HY5的同源基因PuHYH1以及乙烯信號轉(zhuǎn)導途徑的重要轉(zhuǎn)錄因子ERF和乙烯合成的關(guān)鍵基因ACS、ACO,通過對其表達模式分析發(fā)現(xiàn),藍光處理后PuHYH1的表達上調(diào)乙烯合成關(guān)鍵基因PuACS1的表達下調(diào),PuEPF2表達上調(diào)。2.酵母單雜交、EMSA試驗發(fā)現(xiàn)PuHYH1和PnERF2均能夠結(jié)合PuACS1的啟動子;通過GUS報告基因活性測定試驗發(fā)現(xiàn)PuHYH1和PuERF2均可負調(diào)控PuACS1基因的表達。3.通過酵母雙雜交試驗、Pull-down試驗發(fā)現(xiàn)了 PuHYH1與PuERF2之間存在蛋白互作關(guān)系。4.通過GUS報告基因活性測定試驗明確了 PuHYH1與PuERF2蛋白之間的互作可以增強PuERF2對PuACS1的負調(diào)控作用。以上結(jié)果表明:PuHYH1、PuERF2可以直接負調(diào)控PuACS1的表達,PuHYH1與PuERF2互作可以增強PuERF2對PuACS1的負調(diào)控,藍光可能通過以上兩條途徑抑制PuACS1的表達進而抑制乙烯合成。本實驗結(jié)果從分子水平闡明了藍光抑制'南果梨'乙烯合成的機制,對如何提高'南果梨'的貯藏性研究提供了重要的理論基礎(chǔ)。
[Abstract]:The ripening process of fruit is an important factor to determine its storage property. The study of fruit maturation and regulation mechanism has important theoretical and practical significance for artificial regulation of fruit storage. Pyrus ussuriensis Maximis is a unique pear resource in northern China. Most of the autumn pears, including 'Nanguo pear', are respiratory jump type fruits, and their ripening process is mainly regulated by ethylene. In addition, light, temperature and other external environmental factors can also regulate the fruit ripening process. In recent years, many studies have proved that blue light can inhibit ethylene synthesis in the ripening process of respiratory jump fruits, thus affecting the storage ability of fruits, but its molecular mechanism is not clear. In this study, the molecular mechanism of blue light inhibiting the expression of ethylene synthesis key gene PuACS1 and regulating ethylene synthesis during fruit ripening was studied using the fruit of 'Nanguo pear' as the experimental material, and the transcription factor PuHYH1 in its signaling pathway was used to inhibit the expression of the key gene of ethylene synthesis. The main results are as follows: 1. The homologous gene PuHYH1 of important transcription factor HY5 in light signal transduction pathway, the important transcription factor ERF of ethylene signal transduction pathway and the key gene of ethylene biosynthesis, ACSACO, were cloned from the genome of pear. The expression of PuHYH1 upregulated after blue light treatment down-regulated the expression of PuACS1, the key gene of ethylene synthesis, and upregulated the expression of PuEPF2. The results showed that both PuHYH1 and PnERF2 could bind to the promoter of PuACS1, and PuHYH1 and PuERF2 could negatively regulate the expression of PuACS1 gene by GUS reporter gene activity assay. The protein interaction between PuHYH1 and PuERF2 was found by the pull-down test of yeast two-hybrid. 4. GUS reporter gene activity assay showed that the interaction between PuHYH1 and PuERF2 protein could enhance the negative regulation of PuERF2 on PuACS1. These results suggest that the interaction of PuHYH1 and PuERF2 can directly and negatively regulate the expression of PuACS1. The interaction between PuHYH1 and PuERF2 can enhance the negative regulation of PuACS1 by PuERF2. Blue light may inhibit the expression of PuACS1 and inhibit ethylene synthesis through these two pathways. In this study, the mechanism of blue light inhibiting ethylene synthesis of 'Nanguo pear' was elucidated at molecular level, which provided an important theoretical basis for the study on how to improve the storage ability of 'Nanguo pear'.
【學位授予單位】：沈陽農(nóng)業(yè)大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：S661.2
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本文編號：1826477